Tissue engineered blood vessels may potentially revolutionize vascular replacement surgery by providing viable alternatives for often unavailable autologous grafts. Vascular constructs tissue engineered using natural components offer the potential to mimic the responsive nature of native arteries. However in spite of much progress in the field those developed so far have insufficient strength or require an extremely long time of growth in vitro to achieve it. From our own recent efforts to optimize development of vascular grafts tissue engineered using vascular smooth muscle cells (SMC) seeded collagen, we learned that loss of mechanical properties of such constructs is associated with premature scaffold degradation by enzymes released by SMC, called matrix metalloproteinases (MMPs). Blocking MMPs however, proved to also prevent SMC capacity to assemble matrix, process essential for proper construct organization and function. We are proposing to explore a potential construct design improvement, which would preserve the advantageous MMP remodeling functions while preventing premature failure, by providing additional mechanical support during the in vitro assembly through the addition of a biodegradable scaffold. The innovative aspect of this idea is that we seek to identify a scaffold that, in addition to providing mechanical support, will also optimize the characteristics of natural components that have been associated with enhanced mechanical properties of tissue engineered constructs: cell viability and appropriate turn-over and organization of the natural matrix components. For this purpose we will use a novel rational combinatorial approach, which allows testing of a large number of potential chemical and physical characteristics of the biodegradable scaffold. We will also assess whether the synthetic scaffold is susceptible to degradation by MMPs, which would modify its properties and may be related to the body reaction following construct implantation in vivo. For these experiments we will use SMC that we isolate from the arteries of genetically engineered MMP knock-out or transgenic mice, which will allow the specific testing of the role of these enzymes. Our studies should contribute to the improvement of vascular grafts tissue engineered using natural components. We plan to use the gained information to optimize the future design of hybrid grafts containing human vascular smooth muscle and endothelial cells.